Fig. 1: Heavy formaldehyde fixation modulates but does not eliminate chromatin architecture evidence in experimental yeast cultures.

A Pooled occupancy values (FAIRE: blue, MNase: green) compared to gDNA extraction control (purple) with 15 min or 24 h fixation of heat-shocked Saccharomyces cerevisiae. Shading indicates regions with significant peak width shifts (FDR < 0.05). Upstream of highly upregulated GAD1 and HSP26 genes (log2FC = 3.8 and 9.02), changes in occupancy signal morphology are observed (y-axis) relative to the genomic position (x-axis). The 5’ FAIRE peak broadens, while the distinct 5’ MNase nucleosome array transforms into a single peak. B Venn diagrams demonstrate the repeatability of FAIRE (blue) and MNase (green) treatment among technical replicates. Numbers/proportions represent genes with significant peak gain (FDR < 0.05, log10Pval < −6) within 2 kb upstream of the transcription start site (TSS). C Differential DANPOS3 occupancy values were calculated by comparing pooled heat-shocked and optimal growth replicates treated with MNase. Statistical significance was assessed using a two-sided t-test, and the FDR was controlled using the Benjamini–Hochberg procedure to adjust for multiple comparisons. Signal changes are shown for a highly upregulated gene (HSP42, log2FC = 4.86) and a highly downregulated gene (HXT2, log2FC = −2.21) as measured by RNA-Seq in fresh cultures. Shading indicates regions of significant (FDR < 0.05, log10Pval < −6) peak gains (green) or losses (orange). D Total signal log2FC for genes with significant (FDR < 0.05) total peak signal change between pooled replicate heat shock and optimal growth conditions in the 2 kb region upstream of the TSS is plotted against expression log2FC. Genes shared between the 15-min and 24-h time points are shown (green = signal gain; orange = signal loss). Linear regression lines are fitted with correlation coefficients (R) and p-values. E Venn diagram illustrating the overlap between genes identified as upregulated via RNA-Seq and with MNase peak gains in yeast fixed for 24 h. F Gene Ontology (GO) Biological Process enrichment: Genes with significant peak gain across pooled replicates (FDR < 0.05, log10Pval < −6) within 2 kb upstream of the TSS in MNase-treated yeast fixed for 24 h (N = 383) compared to significantly upregulated genes (N = 352) measured by RNA-Seq. GO term enrichment was calculated using Enrichr, with statistical significance assessed using a Fisher exact test. The FDR was controlled using the Benjamini–Hochberg procedure for multiple comparisons. The length of coloured bars corresponds to the Enrichr combined score [log(p-value) * z-score]. MNase GO terms are coloured shades of green, dark blue or grey if they are found in the top 5, top 25 or not within the RNA-Seq GO terms. Source data are provided as a Source Data file.